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. 2003 Oct 1;4(12):1182–1189. doi: 10.1038/sj.embor.7400018

Figure 4.

Figure 4

Phosphoserine 30 modulates AP-1 binding to VAMP4 via PACS-1. (A) AtT20 cells were infected with either wt or HA-PACS-1 vaccinia virus and used for immunoprecipitation with anti-VAMP4 antibody. HA-PACS-1 can be detected in the VAMP4 immunoprecipitates. (B) GST-VAMP4 was incubated with or without CK2, bound to glutathione beads and incubated with PACS-1. PACS-1 binding to wt or the LL/VV VAMP4 is enhanced by the presence of CK2; the S30D mutant shows a CK2-independent binding comparable to phosphorylated VAMP4. The lower panel shows quantitation of the PACS-1 binding to wt VAMP4 and mutants in the presence or absence of CK2. (C) Formation of the ternary VAMP4/PACS-1/AP-1 complex using GST-VAMP4, PACS-1 and purified AP-1. Addition of PACS-1 protein to the VAMP4 LL/VV S30D protein enhances binding of AP-1 compared to VAMP4 LL/VV S30A protein. Quantitation (lower panel) of AP-1 binding to the VAMP4/PACS-1 complex shows a PACS-1-dependent increase.